Both humoral and cellular immune responses are essential components of defenses against pathogenic bacteria, viruses, and parasites. Key players in the immune response, called T lymphocytes, control cellular immunity by virtue of their ability to discriminate between a particular antigen and its close relative. This remarkable antigen specificity of the T lymphocyte responses is due to the presence on the T cell surface of clonally distributed, immunoglobulin-like T cell receptors (TCR) consisting of two non-identical glycosylated polypeptide chains, called α and β. T lymphocytes also express on their surface glycoproteins that are markers of different stages and types of T cell maturation (e.g., T3, T4, and T8 glycoproteins present on CD3+, CD4+ and CD8+ T cells, respectively), which may mediate or augment specific T cell functions.
TCRs interact with antigens that have been processed by the antigen presenting cell (APC) to unfold or cleave the protein into peptide fragments, and are presented on the cell surface as part of a complex with a molecule encoded by genes within the major histocompatibility complex (MHC). CD8+ and CD4+ T lymphocytes interact with peptides bound to the polymorphic region of MHC class I or class II molecules, respectively (Townsend et al., Ann. Rev. Immunol., 7, 601-24 (1989); Rothbard et al., Ann. Rev. Immunol., 9, 527-65 (1991)). The TCR also interacts with proteins that have the capacity to generate intracellular second messenger signals that are essential to triggering T cell activation (Ashwell et al., Ann. Rev. Immunol., 8, 139-67 (1990)), or induction of T cell proliferation or differentiation. For example, recent data show that the CD3 γ, δ, ε: ζ/n complex stably associated with the TCR consists of at least two separate signal transduction modules that initiate second messenger cascades (Letourneur et al., Proc. Natl. Acad. Sci. USA, 88, 8905-09 (1991); Wegener et al., Cell, 68, 83-95 (1992)) which include primary events such as tyrosine phosphorylation (Samelson et al., Cell, 46, 1083-90 (1986)) and secondary events. Such as PIP2 hydrolysis and elevation of [Ca++]i (Weiss et al., Proc. Natl. Acad. Sci. USA, 81, 4169-73 (1984); June et al., J. Immunol., 144, 1591-99 (1990)).
The effect of these TCR-mediated biochemical events on the T cell is influenced by independent receptor-ligand interactions that may generate different types of signals from those evoked by the TCR-ligand interaction (Weaver et al., Proc. Natl. Acad. Sci. USA, 85, 8181-85 (1988); Mueller et al., J. Immunol., 144, 3701-09 (1990); Linsley et al., J. Exp. Med., 173, 721-30 (1991); Koulova et al., J. Exp. Med., 173, 759-62 (1991); Vandenberghe et al., J. Exp. Med., 175, 951-60 (1992)). Studies manipulating the potential of the APC to provide co-stimulation (Quill et al., J. Immunol., 138, 3704-12 (1987); Otten et al., Science, 251, 1228-31 (1991)) have separated effector activities of CD4+ and CD8+ T cells into activities that do (e.g., cytokine interleukin (IL)-2 production) and do not (e.g., cytokine IL-3 production and cell killing) require co-stimulatory signals.
The participation of co-stimulatory signals in control of production of IL-2 (Jenkins et al., Immunol. Rev., 95, 113-35 (1987)), the cytokine that is primarily responsible for clonal expansion following T cell activation, is indicated by the finding that metabolically-inactivated cells bearing ligands for the TCR present on CD4+ cells are unable to effectively stimulate IL-2-dependent T cell proliferation (Bach et al., Immunol. Rev., 35, 76-96 (1977); Germain, J. Immunol., 127, 1964-66 (1981); Jenkins et al., J. Exp. Med., 165, 302-19 (1987)), which suggests there is a critical ‘second signal’ missing in these inactivated cells that operates independently of TCR-regulated second messenger generation or augmentation of TCR occupancy (Mueller et al., J. Immunol., 142, 2617-28 (1989a)). Several candidate receptor-ligand pairs have been suggested for the co-stimulatory pathway, such as the binding of B7 surface protein on the APC by CD28 on the responding T cell (Freeman et al., J. Exp. Med., 174, 625-31 (1991); Gimmi et al., Proc. Natl. Acad. Sci. USA, 88, 6575-79 (1991); Koulova et al., J. Exp. Med., 173, 759-62 (1991); Linsley et al., J. Exp. Med., 173, 721-30 (1991); Reiser et al., Proc. Natl. Acad. Sci. USA, 89, 271-75 (1992); Vandenberghe et al., J. Exp. Med., 175, 951-60 (1992)), and the recognition of the heat-stable antigen on the APC by an uncharacterized T cell counter-receptor (Kay et al., J. Immunol., 145, 1952-59 (1990); Liu et al., J. Exp. Med., 175, 437-45 (1992)).
Activation of the T cell is initiated when some adequate number of TCRs are aggregated at the interface between the T cell and the APC (Singer, Science, 255, 1671-77 (1992); Matis et al., Proc. Natl. Acad. Sci. USA, 80, 6019-23 (1983a); Ashwell et al., J. Immunol., 136, 757-68 (1986)). The extent of receptor-ligand aggregation depends on the number of available receptors on the T cell, the number of available ligands, i.e., peptide-MHC molecule complexes, on the APC, and the affinity of the TCR for the ligand. When a high level of peptide-MHC molecule complexes on the APC fails to induce T cell activation, it is believed this is due to a low affinity of the TCR for the ligand, which prevents receptor occupancy from exceeding the threshold needed for second messenger generation within the T cell (Fiering et al., Genes Dev., 4, 1823-34 (1990).
This affinity-based occupancy model predicts that in the presence of intact, metabolically-active APC capable of delivering co-stimulatory signals, peptide-MHC molecule complexes will be of two types: (1) agonists that can induce full T cell activation, and (2) non-agonists that do not induce T cell activation because of low affinity of the TCR for the peptide-MHC molecule complex, which prevents the number of occupied TCR from reaching the triggering threshold level (Matis et al., Proc. Natl. Acad. Sci. USA, 80, 6019-23 (1983a)). Recently, this model has been challenged by findings showing that substitution of a single residue in the peptide antigen for the TCR on a mouse Th2 clone prevented stimulation of proliferative responses, while permitting IL-4 cytokine production (Evavold et al., Science, 252, 1308-10 (1991)). This indicates that contrary to predictions of the affinity-based occupancy model, certain ligands can stimulate T cell second messenger generation without evoking the full repertoire of effector responses. Moreover, peptide analog-MHC molecule complexes have been described which inhibit the IL-2 response of the T cell response by TCR antagonism, or competition with wild-type ligand for binding to the TCR (De Magistris et al., Cell, 68, 625-34 (1992)). It has been reported that the inhibitory complexes were pure TCR antagonists which lacked capacity to generate intracellular signals (De Magistris et al., Cell, 68, 625-34 (1992)). This finding of an absence of second messenger generation despite fully occupied TCRs is also not predicted by the affinity based occupancy model.
The present invention is predicated on the unexpected discovery that there exist TCR ligands which exhibit selective antagonist properties (referred to herein as “selective antagonists”) and which may also concurrently exhibit agonist properties (referred to herein as mixed “agonists-antagonists”). Specifically, peptide-MHC molecule complexes have been identified which interact with the TCR to actively and selectively inhibit IL-2 production by a mouse T cell clone, without preventing IL-3 production, IL-2Rα upregulation, or cell size enlargement induced by a TCR agonist. Since these new TCR ligands are able to selectively modulate certain T cell effector activities in a TCR-specific manner, they can be considered selective antagonists. These selective antagonists differ from the partial agonists described in Evavold et al., Science, 252, 1308-10 (1991), in that the selective antagonists of the present inventor actively inhibit certain effector responses as opposed to simply failing to stimulate these responses. These selective antagonists differ from the complete antagonists described in De Magistris et al., Cell, 68, 625-34 (1992), in that unlike the complete antagonists, the selective antagonists of the present invention have been shown to selectively inhibit certain effector responses, without affecting others, and act without preventing all T cell signaling.
These results suggest there may be two distinct classes of inhibitory peptide-MHC molecule complexes: selective antagonists and complete antagonists. While members of the latter class would prevent intracellular messenger generation in the T cell by removing TCRs from the functional pool and precluding any effector responses, members of the former class would interfere with certain effector activities based on qualitative differences in requirements for intracellular signalling, possibly, but not necessarily, related to the co-stimulation dependence of the analyzed functions.
The properties of the TCR ligands of the present invention have important implications for models of thymic selection and peripheral T cell activation and provide new pharmacological approaches to the treatment of autoimmune disease, to the problems of graft rejection, and in vaccine design. Moreover, the present invention described herein enables the identification, characterization, development, and utilization of the TCR ligands of the present invention.
Consequently, it is an object of the present invention to provide a TCR ligand which inhibits at least one T cell effector response evoked by fully active peptide-MHC molecule complexes available to responding T cells, without necessarily inhibiting all other effector responses of the T cells. It is a related object of the present invention to provide a TCR ligand which inhibits at least one T cell effector response evoked by fully active peptide-MHC molecule complexes available to responding T cells and which does not substantially inhibit at least one other T cell effector response. It is another object of the present invention to provide a TCR ligand which inhibits co-stimulation dependent T cell effector responses-evoked by fully active peptide-MHC molecule complexes available to responding T cells and which does not block co-stimulation independent T cell effector responses under the same conditions. It is yet another object of the present invention to provide TCR ligands which are selective antagonists and mixed agonists-antagonists. It is a further object of the present invention to provide a method of identifying, as well as preparing, such TCR ligands and of providing improved methods of modulating T cell effector response utilizing such TCR ligands.
These and other objects and advantages of the present invention, as well as additional inventive features, will be apparent from the description of the invention provided herein.